JP5886877B2 - Aloe polysaccharide composition and method - Google Patents

Description

  The present invention relates generally to the field of aloe polysaccharides, and more particularly to compositions of aloe polysaccharides and the use of such compositions as immunomodulators and for the treatment of various types of cancer.

  Without limiting the scope of the invention, its background is described in connection with the composition, method of preparation and therapeutic use of Aloe Vera polysaccharides.

  US Pat. No. 7,196,072 (2007), obtained by Pasco et al., Describes a complex water-soluble polysaccharide fraction with potent immunostimulatory activity isolated from Aloe Vera. The polysaccharide fraction has glucose, galactose, mannose and arabinose as its main components, and has an apparent molecular weight exceeding 2 million daltons. The present invention further describes a pharmaceutical composition comprising the polysaccharide fraction, optionally in combination with acceptable pharmaceutical carriers and / or excipients. These pharmaceutical compositions can be used to provide immune stimulation to such individuals by administering an effective amount of the composition to an individual in need of such treatment.

  US Pat. No. 6,083,508 (2000), obtained by Avalos and Danhof, describes a process for forming an aloe product from only leaf residues obtained after opening aloe leaves and removing internal mesophyll. Is described. According to US Pat. No. 6,083,508, the residue is ground into a slurry and an aloe product is produced from the slurry. In addition, the steps of preparing the aloe product include washing before cutting the aloe leaves, separating the formed slurry into liquid and solid, and further separating the liquid before forming the aloe product. Treatment to remove laxatives. A process including all of the above steps can also be performed to form a liquid.

  US Patent Application No. 2006/0084629 (Needleman and Needleman, 2006) stimulates immune system activity including long, high molecular weight polysaccharides that activate the body's innate immune response to stimulate macrophages, T cells, Disclosed is a combination of two special long chain high molecular weight fractions isolated from Aloe Vera and Maitake TD to trigger increased production of B cells, natural killer cells, cytokines and antibodies. These long-chain polysaccharides, together with other active ingredients, can provide proper immune system support, thereby preventing debilitating diseases such as cancer, heart disease and aging.

US Pat. No. 7,196,072 US Pat. No. 6,083,508 US Patent Application No. 2006/0084629

  The present invention relates to aloe polysaccharide compositions and various types selected as immunomodulators and from among leukemias and lymphomas, prostate cancer, breast cancer and colon cancer, immune diseases, particularly tumors related to immunity. The use of the composition for the treatment of cancer is described.

  The present invention comprises the steps of (i) weighing a predetermined amount of freeze-dried aloe powder, wherein the amount is corrected for moisture content, and (ii) removing the freeze-dried aloe powder. Dissolving in ionized water to form a solution; and (iii) adding an organic solvent to the solution to form a first mixture, wherein the ratio of organic solvent to deionized water is at least 2. A step that is 5: 1; (iv) allowing the first mixture to stand for at least 8 hours; and (v) removing a predetermined volume of supernatant from the first mixture and adding an excess volume of the supernatant solution. Forming a second mixture; (vi) centrifuging the second mixture; (vii) observing the presence of a precipitate in the second mixture; (vii) second Any precipitate in the mixture of If observed, adding an additional amount of organic solvent to the first mixture; and (viii) decanting the supernatant of the first mixture by siphoning, the decantation comprising: Carried out only when no precipitate is observed in the second mixture, (ix) filtering the precipitate from the first mixture using a filter paper and a suction funnel under reduced pressure; x) recovering the polymannan extract powder from the suction funnel by scraping; and (xi) placing the polymannan extract powder in the capped lyophilized flask in a freezer for at least 8 hours. And (xii) lyophilizing the frozen powder of the polymannan extract in a freeze dryer; and (xiii) freezing the polymannan extract. A method for preparing a fine powder of polymannan extract comprising the steps of:

  In one aspect, the method further comprises weighing, labeling and storing the fine powder of polymannan extract in a container. The freeze-dried aloe powder described in the embodiments of the present invention includes Aloe vera, Aloe arborescens, Aloe aristata, Aloe dichotoma, Aloe nyeriensis, Aloe nyeriensis, variegate), derived from an aloe species selected from the group consisting of Aloe barbadensis and Aloe wildii. The freeze-dried aloe powder used in the present invention contains an aloe polysaccharide, which is one or more of a short chain, a medium chain, and a long chain. , Very-large chain polysaccharides, or any combination thereof. In a specific embodiment, the organic solvent is selected from the group consisting of methanol, ethanol, isopropyl alcohol and propanol. In another embodiment, the aloe polysaccharide further comprises a monosaccharide selected from the group consisting of glucose, mannose, arabinose and galactose and has a molecular weight in the range of more than 11,500 daltons to 10,000,000 daltons.

  In another embodiment, the freeze-dried aloe powder has at least 25% aloe polysaccharide. In yet another embodiment, the freeze-dried aloe powder has 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 95% aloe polysaccharide. The freeze-dried aloe powder described in the method of the present invention comprises at least 14% aloe polysaccharide having a molecular weight of 66,000 daltons, at least 9% aloe polysaccharide having a molecular weight of 480,000 daltons, 1 At least 3.5% of aloe polysaccharide having a molecular weight of 1,000,000 daltons and at least 2.4% of aloe polysaccharide having a molecular weight of 2,000,000 daltons.

  In a specific embodiment of the method relating to the aloe polysaccharide composition in freeze-dried aloe powder, about 1.32% to 6.36% of the aloe polysaccharide has a molecular weight of 2,000,000 daltons. 2.55% to 3.89% have a molecular weight of 1,000,000 and 63.85% to 73.36% of the aloe polysaccharide has a molecular weight of 480,000 daltons. In one embodiment, the freeze-dried aloe powder may contain one or more residual low molecular weight species selected from the group consisting of glucose, organic acid, lactic acid, malic acid, citric acid and aspartic acid. In another embodiment, the one or more residual low molecular weight species is present in an amount ranging from 14% to 24%. In yet another embodiment, the fine powder of polymannan extract is used in the treatment of one or more malignacies selected from the group consisting of leukemia and lymphoma, prostate cancer, breast cancer and colon cancer, and Used in the preparation of a pharmaceutical composition to be used for the treatment of one or more immune disorders.

  In another embodiment, the present invention provides a sterile injectable formulation of polymannan extract comprising a predetermined amount of fine polymannan extract dissolved in deionized water and one or more pharmaceutical preservatives. Disclose. One or more pharmaceutical preservatives that can be used in the formulations described above are the group consisting of parabens, benzoic acid and their salts, mercury, quaternary ammonium salts, benzyl alcohol and other related alcohols, and phenol. Selected from. In a specific embodiment, the preservative is benzyl alcohol. In one embodiment, the polymannan extract comprises an aloe polysaccharide, wherein the aloe polysaccharide comprises one or more short chain, medium chain, long chain, very long chain polysaccharides, or any combination thereof. In another embodiment, the aloe polysaccharide further comprises a monosaccharide selected from the group consisting of glucose, mannose, arabinose and galactose.

  In another specific embodiment, the aloe polysaccharide has a molecular weight in the range of more than 11,500 daltons to 10,000,000 daltons, and the polymannan extract has at least 25% aloe polysaccharide, The polymannan extract has 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 95% aloe polysaccharide. In a related embodiment, the polymannan extract comprises at least 14% aloe polysaccharide having a molecular weight of 66,000 daltons, at least 9% has a molecular weight of 480,000 daltons, and at least 3.5% is 1, It has a molecular weight of 000,000 daltons and contains at least 2.4% aloe polysaccharide having a molecular weight of 2,000,000 daltons. The composition of the present invention is used in the treatment of one or more cancers selected from the group consisting of leukemia and lymphoma, prostate cancer, breast cancer and colon cancer, for immunomodulation, immune stimulation, or immune system. It is used for the treatment of individuals with insufficiency or immune disorders. The compositions of the present invention cause a 75-80% increase in one or more natural killer (NK) cells.

  In yet another embodiment, the present invention provides a step of identifying an individual in need of treatment for one or more cancers, and a sterile injectable polymannan extract formulation 2 to 3 times a week, 1 or Injecting at a dosage sufficient to treat two or more cancers, wherein the sterile injectable polymannan extract formulation comprises a predetermined amount of fine polymannan extract dissolved in deionized water and A treatment of one or more cancers selected from the group consisting of leukemia and lymphoma, prostate cancer, breast cancer and colon cancer, comprising a step comprising one or more pharmaceutical preservatives. The method comprises the steps of drawing a blood sample from an individual at one or more predetermined intervals, measuring the level of caspase 3 protein in the blood, and comparing the obtained level with the level prior to injection. And the step of increasing caspase 3 level is directly related to increasing the level of apoptosis of one or more cancer cells.

  In one aspect, the dosage of a sterile injectable polymannan extract formulation depends on the individual's weight, age, ethnicity and gender. In another embodiment, the polymannan extract comprises an aloe polysaccharide, wherein the aloe polysaccharide comprises one or more short chain, medium chain, long chain, very long chain polysaccharides, or any combination thereof. . In yet another embodiment, the aloe polysaccharide has a molecular weight in the range of more than 11,500 daltons to 10,000,000 daltons. The polymannan extract described in the method of the present invention has at least 25% aloe polysaccharide. The polymannan extract of the method of the present invention causes a 75-80% increase in one or more natural killer (NK) cells.

  In one embodiment, the present invention provides: (i) identifying an individual having an immune system deficiency or immune disease and in need of immune modulation or stimulation; and (ii) a predetermined dosage of sterile injectable poly Administering a mannan extract formulation intravenously, wherein the sterile injectable polymannan extract formulation comprises a predetermined amount of a fine polymannan extract dissolved in deionized water and one or more pharmaceutical storages And (iii) withdrawing a blood sample from the individual at one or more predetermined intervals, wherein the dosage is determined by the weight, age, ethnicity and gender of the individual. Measuring the level of tumor necrosis factor-alpha (TNFα) in the blood and comparing the obtained level with the level prior to injection, wherein the increased level of TNFα is And a step of indicating the 疫調 nodes or immunostimulants, discloses a method for immunomodulation or immune stimulation in individuals with immune system deficiency or immune diseases. In a specific embodiment, the immune disease is an immune related tumor. In one embodiment, the polymannan extract comprises an aloe polysaccharide, wherein the aloe polysaccharide comprises one or more short chain, medium chain, long chain, very long chain polysaccharides, or any combination thereof. In another embodiment, the polymannan extract causes a 75-80% increase in one or more natural killer (NK) cells. In yet another embodiment, the polymannan extract has at least 25% aloe polysaccharide.

For a more complete understanding of the features and advantages of the present invention, reference is now made to the detailed description of the invention along with the accompanying figures.
FIG. 4 is a size exclusion chromatogram of aloe polysaccharide showing the retention times of various glucose and mannose subunits. FIG. 4 is a size exclusion chromatogram of Aloe polysaccharide showing peaks corresponding to various glucose and mannose subunits. It is a figure of the proton nuclear magnetic resonance profile of the polymannan extract of this invention. It is a figure of the HPLC chromatogram which shows the quantity of the polysaccharide in each polysaccharide molecule group in the methanol precipitation aloe polysaccharide concentrate. FIG. 5 is a proton nuclear magnetic resonance profile of methanol precipitated aloe polysaccharide concentrate. It is a figure of the HPLC chromatogram which shows the quantity of the polysaccharide in each polysaccharide molecule group in the ethanol precipitation aloe polysaccharide concentrate. FIG. 5 is a proton nuclear magnetic resonance profile of ethanol precipitated aloe polysaccharide concentrate. It is a figure of the HPLC chromatogram which shows the quantity of the polysaccharide in each polysaccharide molecule group in the isopropyl alcohol precipitation aloe polysaccharide concentrate. FIG. 4 is a proton nuclear magnetic resonance profile of isopropyl alcohol precipitated aloe polysaccharide concentrate. It is a figure of the HPLC chromatogram which shows the quantity of the polysaccharide in each polysaccharide molecule group in a propanol precipitation aloe polysaccharide concentrate. FIG. 3 is a proton nuclear magnetic resonance profile of a propanol precipitated aloe polysaccharide concentrate.

  Although the creation and use of various embodiments of the present invention are discussed in detail below, it should be understood that the present invention provides many applicable inventive concepts that can be embodied in a wide variety of specific contexts. is there. The specific embodiments discussed herein are merely illustrative of specific ways to make and use the invention and do not delimit the scope of the invention.

  In order to facilitate understanding of the present invention, a few terms are defined below. Terms defined herein have meanings as commonly understood by a person of ordinary skill in the areas relevant to the present invention. Terms such as “a”, “an”, and “the” are not intended to refer to only a singular component, but are intended to encompass the general types in which an example may be used for illustration. . The terminology herein is used to describe specific embodiments of the invention, but their usage does not delimit the invention, except as explicitly set forth in the claims. Absent.

  The present invention describes a process for preparing polymannan extracts and the use of said extracts in the form of injections as immunostimulatory compounds. Immunostimulation is assessed using human-derived macrophages / monocytes, and cell types are assessed for secretion of tumor necrosis factor alpha (TNFα).

  Aloe polysaccharides are generally considered to be molecules composed mostly of simple sugars, glucose and mannose, having a chain length with a molecular weight of 10,000 Daltons to 10,000,000 Daltons. The higher the mannose content and the longer the chain length, the greater the immunoregulatory activity expressed by the polysaccharide. Various long unbranched chains containing these aloe polysaccharides are listed in Table 1.

  Table 2 lists the aloe polysaccharides having a molecular weight of 100,000 Daltons or more.

  Polymannan extract precursor material is described by the inventors in a prior patent entitled “Method of Processing Aloe Leaves” (US Pat. No. 6,083,508, Avalos and Danhof, 2000). Yes. Certificate of analysis of the polymannan extract precursor material is presented in Table 3.

  FIG. 1 is a size exclusion chromatogram of an aloe polysaccharide preparation showing the retention times of various sizes of glucose and mannose subunits. FIG. 2 is a size exclusion chromatogram of an aloe polysaccharide preparation identifying molecular weights ranging from 100 to 10,000,000 daltons. FIG. 3 is a proton nuclear magnetic resonance profile of the polymannan extract of the present invention. Figure 3 shows (i) standard preservatives-absence of sodium benzoate and potassium sorbate, (ii) presence of a smaller monohexose, (iii) whole leaf methodology in processing raw aloe material. (Iv) malic acid peak-presence of Aloe Vera primary marker, (v) aloelide / acemannan in the polysaccharide portion of the profile supporting the presence of large polysaccharide species The presence of a peak as well as (vi) the presence of an acetyl group that supports the presence of a partially acetylated polysaccharide glucomannan.

Size exclusion chromatography (SEC) of Aloe preparation before polymannan extraction:
Instrument: The HPLC system is a 7250 autosampler paired with a Hitachi L-7100 pump and a Waters 410 differential refractometer. The SEC is a Tosoh Biosep G6000 PWXL TSK Gel 30 cm × 7.8 mm operating at 70 ° C. in a column heater. The molecular weight standards are 2,000,000 Dalton, 1,000,000 Dalton, 480,000 Dalton, 66,000 Dalton and 180 Dalton (glucose) manufactured by Sigma. The mobile phase is deionized water with a flow rate of 0.70 mL / min. The injection volume is 10 uL. The SEC method has been described by Pugh et al. (Pugh N., Ross SA, ElSohly MA, and Pasco, DS (2001). Characterization of Aloeride, a new high-molecular weight polysaccharide from Aloe vera with potent immunomodulatory activity. J Agr Food Chem., 49, 1030-1034).

  Aloe Precipitant Evaluation Study: A 25 ml sample of COATS concentrated aloe was pipetted into a 200 ml beaker and 125 ml of various polysaccharide precipitant solutions were added to the beaker and stirred thoroughly. The precipitated polysaccharide was collected by filtration through tared dehydrated filter paper and after filtration it was placed in a drying oven overnight. The next morning, the dry weight of the precipitated polysaccharide was determined. We studied four alcohol precipitating agents including methanol, ethanol, isopropyl alcohol and propanol. Polysaccharides, wherein various amounts of all molecular species are determined via a powder HPLC procedure, including over 2,000,000 daltons, over 1,000,000 daltons, over 480,000 daltons, over 66,000 daltons Recorded with determination of the amount of polysaccharide in each of the molecular groups and the residual fraction containing very low molecular weight species, such as glucose having a molecular weight of 180 (HPLC data is shown in Tables 6-9). The HPLC chromatograms corresponding to the four precipitants methanol, ethanol, isopropyl alcohol and propanol are shown in FIGS. 4A, 5A, 6A and 7A, respectively. A proton nuclear magnetic resonance profile of the precipitate was also acquired and is shown in FIGS. 4B, 5B, 6B and 7B. The collected data is shown in Table 5.

  A polymannan extract is prepared from the precipitate. The freeze-dried aloe powder described above was weighed after appropriate correction for moisture content. For example, when the moisture content is 3.7% and 80 g is required, the inventors weighed 82.96 g (80 g + (3.7% × 80) g). The weighed aloe powder was completely dissolved in 1 gallon of deionized water (DI) in a stainless steel settling tank. 2.5 gallons of 95% ethanol was added and stirred to ensure complete mixing. The vessel was covered with a stainless steel lid and the mixture was allowed to stand overnight.

  The next day, 2 ml of the clear supernatant was collected, 5 ml of 95% ethanol was added, and the sample was centrifuged at 3000 rpm for 20 minutes. The sample was examined for precipitation and if no precipitate was observed, the precipitation was considered complete. If any significant degree of precipitation was observed, additional 95% ethanol was added to the precipitation tank before continuing. The clear supernatant fluid in the settling tank was decanted by sucking up without disturbing the precipitate at the bottom of the tank. A white funnel at the bottom was separated using a suction funnel (Whatman No. 42® quantitative ashless filter paper). The precipitated material was transferred by scraping into a 600 ml Virtis freeze-dried flask and a thin layer with a large exposed surface area was formed by distributing the material on one side of the flask. The lyophilized flask was placed in a shell freezer overnight. The next day, the chilled lyophilized flask containing the frozen contents was placed in a lyophilizer operating at -90 ° C and 1/3 atm for 24 hours. The lyophizer was turned off and the lyophilized powder was placed in a small powder grinder until it became a uniformly ground fine powder. The ground powder was weighed and placed in a small plastic container, and the container was stored in a freezer.

  Preparation of injectable solution of polymannan extract: Polymannan extract powder (PME) prepared as described above has an aloelide content of at least 2% as measured by size exclusion chromatography. It was weighed (1.5 g) after having been corrected with the water content to have it. 125 ml warming To I water, 1 ml of concentrated HCl was added and stirred, followed by the slow addition of PME powder with constant stirring. Stirring is continued until all the PME powder is dissolved and the solution is clear and colorless, and an additional amount of concentrated HCl is added to obtain a pH of 1.6-1.7 (continuously using a pH meter). Measured). Additional D.D. I water was added to adjust the volume to 150 ml, followed by pH monitoring to ensure a pH of 1.6-1.7. The PME solution was then poured into a Corning® 150 ml filter system flask with a pore size of 0.45 μm. The flask system was placed in a refrigerator and the filtrate was transferred to a Corning® 150 ml filter system flask with a pore size of 0.22 μm and placed in the refrigerator overnight. Under aseptic conditions, the filter top of the filter system was removed and the bottle was sealed with a sterile cap. Them is then transferred to the compounding lab, 0.9% benzyl alcohol is added as a preservative under a sterile hood (because the final product is for multiple dose use), and the solution is placed in a sterile 10 mL glass vial And sealed with a multidosage closure. The vial will be labeled with the name of the physician and patient along with the batch number, control number, date of manufacture, and 6 month expiration date.

  PME immunomodulatory activity assessment: Immunostimulatory activity is assessed using human-derived macrophages / monocytes obtained from the American Type Culture Collection (ATCC), Maryland. Cell type was assessed by secretion of TNFα. A small amount of final PME product was introduced into the culture under standard cellular conditions. Samples were removed at 6, 12 and 24 hours and evaluated for TNFα levels. A specific amount of TNFα was not used due to heterogeneity in different cell batches. In the clinical situation, the immunomodulatory response depends on changing hematological factors such as total white blood cell count, macrophage fraction / monocyte fraction number, number of surface mannose receptors on leukocytes, amount of mannose-binding carrier protein, etc. , Are expected to fluctuate.

  The leukocyte profile fluctuates as cells constantly enter and exit the bloodstream. The affinity of cellular mannose receptors for PME is far greater than that of mannose binding proteins. As new macrophages / monocytes enter the bloodstream, PME is transferred from the circulating mannose binding protein to new cells. PME associated with macrophage / monocyte mannose binding protein results in the release of various cytocommunicators. Cytocommunicators, including TNF-α, IL-1β, INF-γ, IL-2, and IL-6, restore normality to the immune system's impaired surveillance in cancer patients that have lost their tumor detection function. Allows the immune system to identify and remove malignant cells.

  Aloe polysaccharides in polymannan extracts having molecular weights of 1,000,000, 300,000, 100,000, 50,000 and 25,000 all showed caspase activity. This caspase-3, caspase-9 and cytochrome-C activity are key points in the treatment of malignant tumors by the composition of the present invention because caspase-3 is a mediator of tumor cell apoptosis. The immunomodulatory activity of initiator (apical) caspase 3 and effector (executioner) caspase 3 and cytochrome-C has been demonstrated as existing and is a mediator system of tumor cell apoptosis It is regarded.

  We tested the compositions described herein on 104 different types of cancer patients. Leukemia and lymphoma were most responsive to the polymannan extract of the present invention (greater than 98%). Prostate cancer, breast cancer and colon cancer were also responsive to the polymannan extract of the present invention. In the study, polyamman extract was administered as an injection (adminsiteres). 10 mg of polymannan extract was reconstituted in sterile water for injection to give a final concentration of about 10 mg / mL. This was injected 2-3 times a week. Serum samples were then taken from the patients at regular intervals and monitored for caspase 3 activity.

  It is contemplated that any embodiment discussed herein can be implemented with respect to any method, kit, reagent or composition of the invention, and vice versa. Furthermore, the compositions of the invention can be used to achieve the methods of the invention.

  It will be understood that the specific embodiments described herein are shown by way of illustration and not as limitations of the invention. The principal features of this invention can be employed in various embodiments without departing from the scope of the invention. Those skilled in the art will be able to identify many equivalents of the specific procedures described herein without recognizing or using more than routine experimentation. Such equivalents are considered to be within the scope of this invention and are covered by the claims.

  All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which the invention pertains. All publications and patent applications are hereby incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

  The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and / or the specification may mean “1”, but “one or more”, It also coincides with the meanings of “at least one” and “one or more than one”. Although this disclosure supports the definition that the term “or” refers only to alternatives and “and / or”, the term “or” in the claims is expressly directed to refer only to alternatives. Or alternatives are used to mean “and / or” unless the alternative is mutually exclusive. Throughout this application, the term “about” is used to indicate that a value encompasses the inherent variation in device error, the method used to determine the value, or variation that exists between study subjects. Used for.

  As used herein and in the claim (s), the word “comprising” (and any form of comprising such as “comprise” and “comprises”), “having” ( And any form of having such as “have” and “has”), “including” (and any form of inclusion such as “includes” and “include”) or “containing” (And any form of containering such as “contains” and “contain”) are generic or open-ended and do not exclude additional unlisted elements or method steps.

  The term “or combinations thereof” as used herein refers to all permutations and combinations of the listed items preceding the term. For example, “A, B, C, or combinations thereof” means at least one of A, B, C, AB, AC, BC, or ABC, and, in a particular context, if order is important, BA, CA, It is intended to encompass CB, CBA, BCA, ACB, BAC or CAB. Following this example, explicitly included are combinations containing one or more repetitions of one or more items or terms, such as BB, AAA, MB, BBC, AAABCCCC, CBBAAA, CABABB. One skilled in the art will understand that typically there is no limit on the number of items or terms in any combination, except where apparent from the context.

  All of the compositions and / or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. Although the compositions and methods of the present invention have been described in terms of preferred embodiments, they are described in the compositions and / or methods and herein without departing from the concept, spirit and scope of the present invention. It will be apparent to those skilled in the art that variations can be applied in certain method steps or series of steps. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.

(References)
United States Patent No. 7,196,072: High Molecular Weight Polysaccharide Fraction From Aloe Vera with Immunostimulatory Activity
United States Patent No. 6,083,508: Method of Processing Aloe Leaves
United States Patent Application No. 2006/0084629: Immune System Activating Formula Composed of Selected Long Chain Polysaccharides From Natural Sources
¹ Pugh N., Ross SA, ElSohly MA, and Pasco, DS (2001). Characterization of Aloeride, a new high-molecular weight polysaccharide from Aloe vera with potent immunomodulatory activity. J Agr. Food Chem., 49, 1030-1034

Claims (11)

A method for preparing a fine powder of polymannan extract,
Weighing a predetermined amount of freeze-dried aloe powder, wherein the amount is corrected for moisture content;
Dissolving the freeze-dried aloe powder in deionized water to form a solution;
Adding a first volume of organic solvent to the solution to form a first mixture, wherein the ratio of the organic solvent to the deionized water is at least 2.5: 1;
Allowing the first mixture to stand for at least 8 hours;
Extracting a predetermined volume of the supernatant solution from the first mixture and adding a second volume of the organic solvent to the predetermined volume of the supernatant solution to form a second mixture;
Centrifuging the second mixture;
Observing the presence of a precipitate in the second mixture;
Adding a third volume of the organic solvent to the first mixture if any precipitate is observed in the second mixture;
Decanting the supernatant of the first mixture by siphoning, wherein the decantation is performed only if no precipitate is observed in the second mixture;
Filtering the precipitate from the first mixture under reduced pressure using filter paper and a suction funnel;
Recovering the polymannan extract powder from the suction funnel by scraping;
Placing the polymannan extract powder in a capped lyophilized flask in a freezer for at least 8 hours;
Freeze-drying the frozen powder of the polymannan extract in a freeze dryer;
Crushing the lyophilized powder of the polymannan extract into a desired granulate in a crusher.   Claim 1 wherein the freeze-dried aloe powder is derived from an aloe species selected from the group consisting of aloe vera, prickly aloe, ayanishiki, takarokai, aloe nieriensis, aloe variegate, honrokai and aloe wildi. The method described.   The process according to claim 1 or 2, wherein the organic solvent is selected from the group consisting of methanol, ethanol, isopropyl alcohol and propanol.   The freeze-dried aloe powder comprises an aloe polysaccharide, wherein the aloe polysaccharide comprises one or more short chain, medium chain, long chain, very long chain polysaccharides, or any combination thereof. The method in any one of 1-3.   5. The method of claim 4, wherein the aloe polysaccharide has a molecular weight in the range of more than 11,500 daltons to 10,000,000 daltons.   6. A method according to any preceding claim, wherein the freeze-dried aloe powder has at least 25% aloe polysaccharide.   The freeze-dried aloe powder contains at least 14% aloe polysaccharide having a molecular weight of 66,000 daltons, contains at least 9% aloe polysaccharide having a molecular weight of 480,000 daltons, and has a molecular weight of 1,000,000 daltons The method according to any one of claims 1 to 6, comprising at least 3.5% of an aloe polysaccharide having a molecular weight of at least 2.4% of an aloe polysaccharide having a molecular weight of 2,000,000 daltons.   The freeze-dried aloe powder comprises an aloe polysaccharide having a molecular weight of 2,000,000 daltons in the range of 1.32% to 6.36% and having a molecular weight of 1,000,000 daltons In an amount of 2.55% to 3.89% and an aloe polysaccharide having a molecular weight of 480,000 daltons in the range of 63.85% to 73.36%. The method of crab.   One or more freeze-dried aloe powder selected from the group consisting of monosaccharides, organic acids, lactic acid, malic acid, citric acid and aspartic acid selected from the group consisting of glucose, mannose, arabinose, and galactose 9. A process according to any of claims 1 to 8, which may contain residual low molecular weight species.   10. The method of claim 9, wherein the one or more residual low molecular weight species is present in an amount ranging from 14% to 24%.   The fine powder of polymannan extract is used in the treatment of one or more malignant tumors selected from the group consisting of leukemia, lymphoma, prostate cancer, breast cancer and colon cancer, and in the treatment of one or more immune disorders. 11. A pharmaceutical composition used for or for the preparation of a pharmaceutical composition that causes a 75-80% increase in one or more natural killer (NK) cells. The method described.